In nature, proteins are assembled into sophisticated and exceedingly ordered structures that allow them to perform numerous functions supporting varying forms of life. The exquisite design of natural proteins prompted researchers to exploit it in synthetic biology to engineer molecules that can self-regulate into nanoparticles with desired structure and that may be employed for numerous purposes like storage of gas, enzyme catalysts, drug delivery, intracellular drug and more.
Cytoplasmic polyhedrosis viruses infecting insects are embedded in protein crystals known as polyhedra that shield the virus from damage. The structure of polyhedra crystals suggests that they can cater as robust containers that can incorporate and protect foreign molecules from degradation, ensuring their compositional and functional stability.
Extreme stability of polyhedral under rigid conditions is offered by dense packing of polyhedirn monomers in crystals with solvent channels of highly low porosity that however limits the incorporation of foreign particles. Study group headed by Satoshi Abe and Takafumi Ueno at Tokyo Institute of Technology hypothesized that is a porous framework inside the PhCs is extended without compromising crystal stability, PhCs can be employed for storage and accumulation of exogenous molecules in living cells.
As in natural PhCs, polyhedirn monomers create a trimer, the researchers assumed that if amino acid residues at the contact interface of each trimer are deleted then the porosity of the leading crystals would be enhanced. To accomplish this goal, they genetically engineered polyhedirn monomers that were then expressed and self-organized in Spodoptera frugiperda IPLB-Sf21AE, the larva of an armyworm moth, infected with baculovirus.
The mutant PhCs maintained crystal lattice of the wild-sort PhC but had drastically extended porosity due to the deletion of amino acid residues with the rearrangement of inter- and intra-molecular hydrogen bonds. As a result, the structured crystals could absorb 2-4 times more exogenous molecules compared to the wild sort PhC, with up to 5,000 fold condensation of the dyes from the 10 uM solution..
As a further step, the researchers examined the performance of the mutant crystals in living insect cells. PhCs revealed high stability in the intracellular environment. Most importantly, the mutant crystals could gather and retain the dyes in living cells, while the natural crystals could not.
Rationale crystal design employed by researchers at the Tokyo institute of Technology offers a strong tool for structural manipulation of self-organized protein crystals to obtain porous nanomaterials with regulated absorption properties.
The engineered porous PhCs can be employed as protein containers for in vivo crystal structure analysis of the cellular molecules and bioorthogonal chemistry in numerous sorts of living cells. Since small crystals with just a few microns size were obtained, the structure analyses were performed at beamlines BL32XU and BL41XU
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